scholarly journals Microtubule-mitochondrial attachment determines cell division symmetry and polarity in fission yeast

2021 ◽  
Author(s):  
Leeba Ann Chacko ◽  
Vaishnavi Ananthanarayanan
Keyword(s):  
Cell Division ◽  
Fission Yeast ◽  
Daughter Cell ◽  
Significant Proportion ◽  
Intrinsic Factor ◽  
The Other ◽  
Wild Type ◽  
Feedback Controls ◽  
Unequal Distribution ◽  
Mutant Cells

Association with microtubules inhibits the fission of mitochondria in Schizosachharomyces pombe. Here we show that this attachment of mitochondria to microtubules is an important cell intrinsic factor in determining division symmetry as well as maintaining polarity. By comparing mutant cells that exhibited enhanced attachment and no attachment of mitochondria to microtubules (Dnm1Δ and Mmb1Δ respectively), we show that microtubules in these mutants displayed aberrant dynamics compared to wild-type cells, which resulted in errors in nuclear positioning. This translated to cell division asymmetry in a significant proportion of both Dnm1Δ and Mmb1Δ cells. So too, microtubule pivoting was enhanced in both mitochondrial mutants, resulting in a fraction of the cells in these populations displaying polarity defects. The asymmetric division in Dnm1Δ and Mmb1Δ cells resulted in unequal distribution of mitochondria, with the daughter cell that received more mitochondria growing faster than the other daughter. Taken together, we show the existence of homeostatic feedback controls between mitochondria and microtubules in fission yeast, which directly influence mitochondrial partitioning and thereby, cell growth.

scholarly journals The UDP-Glc:Glycoprotein Glucosyltransferase Is Essential for Schizosaccharomyces pombe Viability under Conditions of Extreme Endoplasmic Reticulum Stress

1998 ◽  
Vol 143 (3) ◽  
pp. 625-635 ◽  
Author(s):  
Sandra Fanchiotti ◽  
Fabiana Fernández ◽  
Cecilia D'Alessio ◽  
Armando J. Parodi
Keyword(s):  
Cell Viability ◽  
Schizosaccharomyces Pombe ◽  
Double Mutant ◽  
Expression Vector ◽  
The Other ◽  
Wild Type ◽  
Glucosidase Ii ◽  
Wild Type Phenotype ◽  
Polypeptide Chains ◽  
Mutant Cells

Interaction of monoglucosylated oligosaccharides with ER lectins (calnexin and/or calreticulin) facilitates glycoprotein folding but this interaction is not essential for cell viability under normal conditions. We obtained two distinct single Schizosaccharomyces pombe mutants deficient in either one of the two pathways leading to the formation of monoglucosylated oligosaccharides. The alg6 mutant does not glucosy- late lipid-linked oligosaccharides and transfers Man9GlcNAc2 to nascent polypeptide chains and the gpt1 mutant lacks UDP-Glc:glycoprotein glucosyltransferase (GT). Both single mutants grew normally at 28°C. On the other hand, gpt1/alg6 double-mutant cells grew very slowly and with a rounded morphology at 28°C and did not grow at 37°C. The wild-type phenotype was restored by transfection of the double mutant with a GT-encoding expression vector or by addition of 1 M sorbitol to the medium, indicating that the double mutant is affected in cell wall formation. It is suggested that facilitation of glycoprotein folding mediated by the interaction of monoglucosylated oligosaccharides with calnexin is essential for cell viability under conditions of extreme ER stress such as underglycosylation of proteins caused by the alg6 mutation and high temperature. In contrast, gls2/alg6 double-mutant cells that transfer Man9GlcNAc2 and that are unable to remove the glucose units added by GT as they lack glucosidase II (GII), grew at 37°C and had, when grown at 28°C, a phenotype of growth and morphology almost identical to that of wild-type cells. These results indicate that facilitation of glycoprotein folding mediated by the interaction of calnexin and monoglucosylated oligosaccharides does not necessarily require cycles of reglucosylation–deglucosylation catalyzed by GT and GII.

scholarly journals The number of cytokinesis nodes in mitotic fission yeast scales with cell volume

2021 ◽  
Author(s):  
Wasim A Sayyad ◽  
Thomas D Pollard
Keyword(s):  
Plasma Membrane ◽  
Fission Yeast ◽  
Large Population ◽  
Yeast Cells ◽  
Wild Type ◽  
Contractile Ring ◽  
Node Number ◽  
Nmt1 Promoter ◽  
Wide Range ◽  
Mutant Cells

Cytokinesis nodes are assemblies of stoichiometric ratios of proteins associated with the plasma membrane, which serve as precursors for the contractile ring during cytokinesis by fission yeast. The total number of nodes is uncertain, because of the limitations of the methods used previously. Here we used the ~140 nm resolution of Airyscan confocal microscopy to resolve a large population of dim, unitary cytokinesis nodes in 3D reconstructions of whole fission yeast cells. Wild-type fission yeast cells make about 200 unitary cytokinesis nodes. Most, but not all of these nodes condense into a contractile ring. The number of cytokinesis nodes scales with cell size in four strains tested, although wide rga4Δ mutant cells form somewhat fewer cytokinesis nodes than expected from the overall trend. The surface density of Pom1 kinase on the plasma membrane around the equators of cells is similar with a wide range of node numbers, so Pom1 does not control cytokinesis node number. However, varying protein concentrations with the nmt1 promoter showed that the numbers of nodes increase above a baseline of about 200 with the total cellular concentration of either Pom1 or the kinase Cdr2.

scholarly journals Most Retroviral Recombinations Occur during Minus-Strand DNA Synthesis

2000 ◽  
Vol 74 (5) ◽  
pp. 2313-2322 ◽  
Author(s):  
Jiayou Zhang ◽  
Ling-Yun Tang ◽  
Ting Li ◽  
Yan Ma ◽  
Christy M. Sapp
Keyword(s):  
Dna Synthesis ◽  
Daughter Cell ◽  
Frameshift Mutation ◽  
The Other ◽  
Minus Strand ◽  
Wild Type ◽  
Chromosomal Dna ◽  
Rna Molecules ◽  
Double Stranded Dna ◽  
Frameshift Mutant

ABSTRACT Retroviral RNA molecules are plus, or sense in polarity, equivalent to mRNA. During reverse transcription, the first strand of the DNA molecule synthesized is minus-strand DNA. After the minus strand is polymerized, the plus-strand DNA is synthesized using the minus-strand DNA as the template. In this study, a helper cell line that contains two proviruses with two different mutated gfp genes was constructed. Recombination between the two frameshift mutant genes resulted in a functional gfp. If recombination occurs during minus-strand DNA synthesis, the plus-strand DNA will also contain the functional sequence. After the cell divides, all of its offspring will be green. However, if recombination occurs during plus-strand DNA synthesis, then only the plus-strand DNA will contain the wild-type gfp sequence and the minus-strand DNA will still carry the frameshift mutation. The double-stranded DNA containing this mismatch was subsequently integrated into the host chromosomal DNA of D17 cells, which were unable to repair the majority of mismatches within the retroviral double-strand DNA. After the cell divided, one daughter cell contained the wild-type gfp sequence and the other daughter cell contained the frameshift mutation in thegfp sequence. Under fluorescence microscopy, half the cells in the offspring were green and the other half of the cells were colorless or clear. Thus, we demonstrated that more than 98%, if not all, retroviral recombinations occurred during minus-strand DNA synthesis.

scholarly journals p63cdc13, a B-type cyclin, is associated with both the nucleolar and chromatin domains of the fission yeast nucleus.

1993 ◽  
Vol 4 (11) ◽  
pp. 1087-1096 ◽  
Author(s):  
I M Gallagher ◽  
C E Alfa ◽  
J S Hyams
Keyword(s):  
Electron Microscopy ◽  
Fission Yeast ◽  
The Other ◽  
Cyclin B ◽  
Immunogold Electron Microscopy ◽  
Wild Type ◽  
Gold Particles ◽  
Phase Protein ◽  
M Phase ◽  
Mitotic Cyclin

The cellular distribution of the fission yeast mitotic cyclin B, p63cdc13, was investigated by a combination of indirect immunofluorescence light microscopy, immunogold electron microscopy, and nuclear isolation and fractionation. Immunofluorescence microscopy of wild-type cells and the cold-sensitive mutant dis2.11 with a monospecific anti-p63cdc13 antiserum was consistent with the association of a major subpopulation of fission yeast M-phase protein kinase with the nucleolus. Immunogold electron microscopy of freeze-substituted wild-type cells identified two nuclear populations of p63cdc13, one associated with the nucleolus, the other with the chromatin domain. To investigate the cell cycle regulation of nuclear labeling, the mutant cdc25.22 was synchronized through mitosis by temperature arrest and release. Immunogold labeling of cells arrested at G2M revealed gold particles present abundantly over the nucleolus and less densely over the chromatin region of the nucleus. Small vesicles around the nucleus were also labeled by anti-p63cdc13, but few gold particles were detected over the cytoplasm. Labeling of all cell compartments declined to zero through mitosis. Cell fractionation confirmed that p63cdc13 was substantially enriched in both isolated nuclei and in a fraction containing small vesicles and organelles. p63cdc13 was not extracted from nuclei by treatment with RNase A, Nonidet P40 (NP-40), Triton X-100, and 0.1 M NaCl, although partial solubilization was observed with DNase I and 1 M NaCl. A known nucleolar protein NOP1, partitioned in a similar manner to p63cdc13, as did p34cdc2, the other subunit of the M-phase protein kinase. We conclude that a major subpopulation of the fission yeast mitotic cyclin B is targeted to structural elements of the nucleus and nucleolus.

Protein transport and flagellum assembly dynamics revealed by analysis of the paralysed trypanosome mutant snl-1

1999 ◽  
Vol 112 (21) ◽  
pp. 3769-3777 ◽  
Author(s):  
P. Bastin ◽  
T.J. Pullen ◽  
T. Sherwin ◽  
K. Gull
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Trypanosoma Brucei ◽  
Daughter Cell ◽  
Protein Transport ◽  
Retrograde Transport ◽  
Transport Systems ◽  
Major Constituent ◽  
Wild Type ◽  
Detergent Extraction

The paraflagellar rod (PFR) of Trypanosoma brucei is a large, complex, intraflagellar structure that represents an excellent system in which to study flagellum assembly. Molecular ablation of one of its major constituents, the PFRA protein, in the snl-1 mutant causes considerable alteration of the PFR structure, leading to cell paralysis. Mutant trypanosomes sedimented to the bottom of the flask rather than staying in suspension but divided at a rate close to that of wild-type cells. This phenotype was complemented by transformation of snl-1 with a plasmid overexpressing an epitope-tagged copy of the PFRA gene. In the snl-1 mutant, other PFR proteins such as the second major constituent, PFRC, accumulated at the distal tip of the growing flagellum, forming a large dilation or ‘blob’. This was not assembled as filaments and was removed by detergent-extraction. Axonemal growth and structure was unmodified in the snl-1 mutant and the blob was present only at the tip of the new flagellum. Strikingly, the blob of unassembled material was shifted towards the base of the flagellum after cell division and was not detectable when the daughter cell started to produce a new flagellum in the next cell cycle. The dynamics of blob formation and regression are likely indicators of anterograde and retrograde transport systems operating in the flagellum. In this respect, the accumulation of unassembled PFR precursors in the flagellum shows interesting similarities with axonemal mutants in other systems, illustrating transport of components of a flagellar structure during both flagellum assembly and maintenance. Observation of PFR components indicate that these are likely to be regulated and modulated throughout the cell cycle.

scholarly journals Cell Structure Changes in the Hyperthermophilic Crenarchaeon Sulfolobus islandicus Lacking the S-Layer

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Changyi Zhang ◽  
Rebecca L. Wipfler ◽  
Yuan Li ◽  
Zhiyu Wang ◽  
Emily N. Hallett ◽  
...  
Keyword(s):  
Cell Division ◽  
Cell Size ◽  
Cell Fusion ◽  
Cell Biology ◽  
Cell Structure ◽  
Evolutionary Relationship ◽  
Wild Type ◽  
Sulfolobus Islandicus ◽  
Mutant Cells

ABSTRACT Rediscovery of the ancient evolutionary relationship between archaea and eukaryotes has revitalized interest in archaeal cell biology. Key to the understanding of archaeal cells is the surface layer (S-layer), which is commonly found in Archaea but whose in vivo function is unknown. Here, we investigate the architecture and cellular roles of the S-layer in the hyperthermophilic crenarchaeon Sulfolobus islandicus. Electron micrographs of mutant cells lacking slaA or both slaA and slaB confirm the absence of the outermost layer (SlaA), whereas cells with intact or partially or completely detached SlaA are observed for the ΔslaB mutant. We experimentally identify a novel S-layer-associated protein, M164_1049, which does not functionally replace its homolog SlaB but likely assists SlaB to stabilize SlaA. Mutants deficient in the SlaA outer layer form large cell aggregates, and individual cell size varies, increasing significantly up to six times the diameter of wild-type cells. We show that the ΔslaA mutant cells exhibit more sensitivity to hyperosmotic stress but are not reduced to wild-type cell size. The ΔslaA mutant contains aberrant chromosome copy numbers not seen in wild-type cells, in which the cell cycle is tightly regulated. Together, these data suggest that the lack of SlaA results in either cell fusion or irregularities in cell division. Our studies show the key physiological and cellular functions of the S-layer in this archaeal cell. IMPORTANCE The S-layer is considered to be the sole component of the cell wall in Sulfolobales, a taxonomic group within the Crenarchaeota whose cellular features have been suggested to have a close relationship to the last archaea-eukaryote common ancestor. In this study, we genetically dissect how the two previously characterized S-layer genes as well as a newly identified S-layer-associated protein-encoding gene contribute to the S-layer architecture in Sulfolobus. We provide genetic evidence for the first time showing that the slaA gene is a key cell morphology determinant and may play a role in Sulfolobus cell division or/and cell fusion.

scholarly journals Fission yeast minichromosome loss mutants mis cause lethal aneuploidy and replication abnormality.

1994 ◽  
Vol 5 (10) ◽  
pp. 1145-1158 ◽  
Author(s):  
K Takahashi ◽  
H Yamada ◽  
M Yanagida
Keyword(s):  
Cell Division ◽  
Fission Yeast ◽  
Rna Splicing ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Unequal Distribution ◽  
High Loss ◽  
Colony Color ◽  
Cell Cycle Block ◽  
Yeast Mutants

Precise chromosome transmission in cell division cycle is maintained by a number of genes. The attempt made in the present study was to isolate temperature-sensitive (ts) fission yeast mutants that display high loss rates of minichromosomes at permissive or semipermissive temperature (designated mis). By colony color assay of 539 ts strains that contain a minichromosome, we have identified 12 genetic loci (mis1-mis12) and determined their phenotypes at restrictive temperature. Seven of them are related to cell cycle block phenotype at restrictive temperature, three of them in mitosis. Unequal distribution of regular chromosomes in the daughters is extensive in mis6 and mis12. Cells become inviable after rounds of cell division due to missegregation. The phenotype of mis5 is DNA replication defect and hypersensitivity to UV ray and hydroxyurea. mis5+ encodes a novel member of the ubiquitous MCM family required for the onset of replication. The mis5+ gene is essential for viability and functionally distinct from other previously identified members in fission yeast, cdc21+, nda1+, and nda4+. The mis11 mutant phenotype was the cell division block with reduced cell size. Progression of the G1 and G2 phases is blocked in mis11. The cloned mis11+ gene is identical to prp2+, which is essential for RNA splicing and similar to a mammalian splicing factor U2AF65.

scholarly journals NuMA is required for the proper completion of mitosis.

1993 ◽  
Vol 120 (4) ◽  
pp. 947-957 ◽  
Author(s):  
D A Compton ◽  
D W Cleveland
Keyword(s):  
Daughter Cell ◽  
Coiled Coil ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Tail Domain ◽  
Wild Type ◽  
Dominant Phenotype ◽  
Chromosome Separation ◽  
Mutant Cells ◽  
Globular Head

NuMA is a 236-kD intranuclear protein that during mitosis is distributed into each daughter cell by association with the pericentrosomal domain of the spindle apparatus. The NuMA polypeptide consists of globular head and tail domains separated by a discontinuous 1500 amino acid coiled-coil spacer. Expression of human NuMA lacking its globular head domain results in cells that fail to undergo cytokinesis and assemble multiple small nuclei (micronuclei) in the subsequent interphase despite the appropriate localization of the truncated NuMA to both the nucleus and spindle poles. This dominant phenotype is morphologically identical to that of the tsBN2 cell line that carries a temperature-sensitive mutation in the chromatin-binding protein RCC1. At the restrictive temperature, these cells end mitosis without completing cytokinesis followed by micronucleation in the subsequent interphase. We demonstrate that the wild-type NuMA is degraded in the latest mitotic stages in these mutant cells and that NuMA is excluded from the micronuclei that assemble post-mitotically. Elevation of NuMA levels in these mutant cells by forcing the expression of wild-type NuMA is sufficient to restore post-mitotic assembly of a single normal-sized nucleus. Expression of human NuMA lacking its globular tail domain results in NuMA that fails both to target to interphase nuclei and to bind to the mitotic spindle. In the presence of this mutant, cells transit through mitosis normally, but assemble micronuclei in each daughter cell. The sum of these findings demonstrate that NuMA function is required during mitosis for the terminal phases of chromosome separation and/or nuclear reassembly.

scholarly journals Mps1 expression is critical for chromosome orientation and segregation in yeast with high ploidy

2020 ◽  
Author(s):  
Ashlea Sartin ◽  
Madeline Gish ◽  
Jillian Harsha ◽  
Dawson Haworth ◽  
Rebecca LaVictoire ◽  
...  
Keyword(s):  
Cell Division ◽  
Chromosome Number ◽  
Cancer Cells ◽  
Chromosome Segregation ◽  
Daughter Cell ◽  
Yeast Cells ◽  
Loss Of Function ◽  
Master Regulator ◽  
High Ploidy ◽  
Mutant Cells

AbstractIn aneuploid cancer cells, the chromosome segregation apparatus is sensitive to increased chromosome number. The conserved protein kinase, Mps1, is a critical actor of this machinery, orienting the chromosomes properly on the spindle. Abnormally high levels of this kinase have been found in tumors with elevated chromosome number. However, it remains unclear, mechanistically, if and how cells with higher ploidy become dependent upon increased Mps1 levels. To answer these questions, we explored Mps1 dependence in yeast cells with increased sets of chromosomes. We discovered that having more chromosomes affects the ability of cells to orient chromosomes properly. The cells with increased numbers of chromosomes are particularly sensitive to the reduction of Mps1 activity. In mps1 loss of function mutants, cells display an extended prometaphase with a longer spindle and a delay in orienting properly the chromosomes. Altogether, our results suggest that increased numbers of chromosomes render cells more dependent on Mps1 for orienting chromosomes on the spindle. The phenomenon described here may be relevant in understanding why hyperdiploid cancer cells become excessively reliant on high Mps1 expression for successful chromosome segregation.Author summaryMost cells in solid tumors usually carry far more chromosomes than normal cells. Losing or gaining chromosomes during cell division can lead to aneuploidy (an abnormal number of chromosomes), cancer, and other diseases. Mps1 is a master regulator of cell division that is critical to keep the correct number chromosomes in each daughter cell. This master regulator has been shown to target and affect the function of various actors involved in cell division. Abnormally high levels of this master regulator are found in tumors with elevated chromosome numbers. The high levels of this regulator appear to be protecting these tumor cells. To answer if and how cells with higher ploidy become so dependent of Mps1, we generated yeast cells with increased set of chromosomes. Here, we report that cells with elevated chromosome number are particularly sensitive to the reduction of Mps1 level. In cells with higher ploidy and reduced level of Mps1, the progression during cell division is delayed. In the mutant cells, their ability to properly orient and segregate their chromosomes on the spindle is greatly reduced.

scholarly journals The phosphatase inhibitor Sds23 regulates cell division symmetry in fission yeast

2019 ◽  
Author(s):  
Katherine L. Schutt ◽  
James B. Moseley
Keyword(s):  
Cell Division ◽  
Fission Yeast ◽  
Cell Polarity ◽  
Protein Phosphatase ◽  
Yeast Cells ◽  
Spatial Control ◽  
Anchoring Proteins ◽  
Family Protein ◽  
Mutant Cells ◽  
Assembly Site

AbstractAnimal and fungal cells divide through the assembly, anchoring, and constriction of a contractile actomyosin ring (CAR) during cytokinesis. The timing and position of the CAR must be tightly controlled to prevent defects in cell division, but many of the underlying signaling events remain unknown. The conserved heterotrimeric protein phosphatase PP2A controls the timing of events in mitosis, and upstream pathways including Greatwall-Ensa regulate PP2A activity. A role for PP2A in CAR regulation has been less clear, although loss of PP2A in yeast causes defects in cytokinesis. Here, we report that Sds23, an inhibitor of PP2A family protein phosphatases, promotes the symmetric division of fission yeast cells through spatial control of cytokinesis. We found that sds23Δ cells divide asymmetrically due to misplaced CAR assembly, followed by sliding of the CAR away from its assembly site. These mutant cells exhibit delayed recruitment of putative CAR anchoring proteins including the glucan synthase Bgs1. Our observations likely reflect a broader role for regulation of PP2A in cell polarity and cytokinesis because sds23Δ phenotypes were exacerbated when combined with mutations in the fission yeast Ensa homolog, Igo1. These results identify the PP2A regulatory network as a critical component in the signaling pathways coordinating cytokinesis.

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